[Survival of patients with primary central nervous system diffuse large B-cell lymphoma: impact of gene aberrations and protein overexpression of bcl-2 and C-MYC, and selection of chemotherapy regimens].

Objective: To investigate the impact of clinicopathological features, gene rearrangements and protein expression of bcl-6, bcl-2, C-MYC and chemotherapy regime on the prognosis of patients with primary central nervous system diffuse large B-cell lymphoma (PCNS-DLBCL). Methods: Thirty-three cases of PCNS-DLBCL diagnosed from January 2006 to December 2016 at Zhejiang Cancer Hospital were collected. The expression of CD10, bcl-6, bcl-2, MUM1 and MYC were detected by immunohistochemical staining (IHC). The presence of EB virus was detected by in situ hybridization(EBER). Copy number variation (ICN) and translocation status of bcl-6, bcl-2 and C-MYC genes were detected by fluorescence in situ hybridization (FISH). The relationship between the above indexes and the prognosis was analyzed by univariate, bivariate survival analysis and multiple Cox hazard regression analysis. Results: The study included 33 patients of PCNS-DLBCL, without evidence of primary or secondary immunodeficient disease. Male to female ratio was 1.36∶1.00, and the average age was 56 years. Twenty cases had single lesion while 13 had multiple lesions. Deep brain involvement was seen in 12 cases. All patients underwent partial or total tumor resection. Five patients received whole brain post-surgery radiotherapy, nine patients received high-dose methotrexate (HD-MTX) based chemotherapy, and 12 patients received whole-brain radiotherapy combined with HD-MTX based chemotherapy. Severn patients received no further treatment and rituximab was used in 8 patients. According to the Hans model, 27 cases were classified as non-GCB subtypes (81.8%). Bcl-2 was positive in 25 cases (75.8%, 25/33) and highly expressed in 8 (24.2%). MYC was positive in 12 cases (36.4%) and double expression of bcl-2 and MYC was seen in 6 cases. EBER positive rate was 10.0%(3/30), all of which had multiple lesions. Two bcl-6 gene translocations and 3 amplifications were found in 28 patients. Two translocations, 3 ICN or with both bcl-2 gene translocation and ICN were found in 30 patients. Four ICNs of C-MYC gene were found in 28 patients. Elevated protein in cerebrospinal fluid (CSF) was found in 13 patients. LDH increased in 10 cases. Follow-up period was 2-90 months with the average survival time of (23.0±3.7) months and two-year survival rate of 39.0%. Univariate survival analysis showed that overexpression of bcl-2 protein (≥70%) and MYC protein (≥40%), bcl-2 gene abnormality (including copy number increase and translocation), C-MYC gene copy number increased were adverse factors for survival. C-MYC/ bcl-2 gene double hit was seen in 2 cases. Bivariate survival analysis found that of bcl-2/MYC protein double expression and bcl-2 and C-MYC genes double aberration were significantly associated with adverse outcomes. Cox multivariate risk regression analysis found that gender, cerebrospinal fluid protein increasing, and ICN of C-MYC gene were independent poor prognostic factors. DH-MTX based comprehensive chemotherapy was associated with better prognosis. Conclusions: Double hit at genomic level (copy number variations and gene rearrangements) and double protein expression of bcl-2 and C-MYC in PCNS-DLBCL are significantly associated with an adverse outcome. DH-MTX based comprehensive treatment may prolong the patient survival.

[Changes of resistant phenotype and CRISPR/Cas system of four Shigella strains passaged for 90 times without antibiotics].

Objective: To explore the stability of resistant phenotypes and changes of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) gene system on four Shigella strains in the absence of antibiotics. Methods: Four clinical isolated Shigella strains that resistant to different antibiotics were consecutive passaged for 90 times without antibiotics. Agar dilution method was used to determine the minimum inhibitory concentration of Shigella strains. After sequence analysis with PCR, CRISPR Finder and Clustal X 2.1 were applied to identify the changes of CRISPR loci in the Shigella strains. Results: After the consecutive transfer of 90 generations, sensitivity to certain antibiotics of four Shigella strains with different drug resistant spectrums increased. Mel-sf1998024/zz resistance to ampicillin, cephalexin, cefotaxime, chloramphenicol decreased, mel-s2014026/sx resistance to norfloxacin, trimethoprim decreased, mel-sf2004004/sx drug resistance to ampicillin, cefuroxime, cefotaxime, chloramphenicol, trimethoprim decreased and mel-sf2013004/bj resistance to chloramphenicol decreased. The spacer of which matched gene codes Cas and its upstream repeat in 3'end of CRISPR3 got lost in mel-sf1998024/zz and mel-sf2013004/bj. Conclusions:Shigella strains could reduce or lose their resistance to some antibiotics after consecutive transfers, without the interference of antibiotics. CRISPR3 locus had dynamic spacers in Shigella strains while CRISPR3 locus and cas genes might have been co-evolved.

Kinetics of aristolochic acid I after oral administration of Radix Aristolochiae or Guanxinsuhe preparation in canines.

ETHNOPHARMACOLOGICAL RELEVANCE: Aristolochic acid I (AAI), a major component derived from Aristolochia species, which have been known for a long time and remain in use today, particularly in Asia and Central America. It has been confirmed to induce a type of so-called aristolochic acid nephropathy (AAN) and involved in the development of Balkan endemic nephropathy (BEN). AIM OF THE STUDY: To investigate the kinetic of AAI in beagle dogs after single-dose oral administration of Radix Aristolochiae or its preparation, Guanxinsuhe, as well as the effects of compound compatibility in traditional Chinese medicine on the pathologic processes of AAN. MATERIALS AND METHODS: Beagle dogs were orally administrated Radix Aristolochiae (0.3 g/kg/day), Guanxinsuhe preparation (0.9 g/kg/day) (with an identical dosage of AAI), and empty capsules respectively for 180 days. Canines (n=2) were euthanized on day 90, 180, 210, HPLC was established to determine the AAI level in plasma and the kinetic behaviors of AAI in dogs were elucidated after single dosing. Hematoxylin and eosin (H&E)-staining was applied for histopathologic examination to evaluate the pathological status of kidneys. RESULTS: Compared to canines with Radix Aristolochiae treatment, the Cmax, AUC, Tmax, and t(1/2ß) of AAI in Guanxinsuhe preparation group were elevated, while t(1/2α) of AAI was decreased. The results indicated the co-existing components in Guanxinsuhe preparation could increase the absorption, accelerate the distribution, but delay the absorption and elimination of AAI. After long-term dosing, animals treated with Radix Aristolochiae were found with more severe renal impairment and higher AAI level in plasma. CONCLUSIONS: It was demonstrated that the compound compatibility in Guanxinsuhe preparation can affect the kinetic process of AAI and attenuate the toxic effect on kidney when the duration of treatment was prolonged.

Ultraviolet electroluminescence from randomly assembled n-SnO(2) nanowiresp-GaN:Mg heterojunction.

Electroluminescence characteristics of a heterojunction light-emitting diode, which was fabricated by depositing a layer of randomly assembled n-SnO(2) nanowires on p-GaN:Mg/sapphire substrate via vapor transport method, were investigated at room temperature. Peak wavelength emission at around 388 nm was observed for the diode under forward bias. This is mainly related to the radiative recombination of weakly bounded excitons at the shallow-trapped states of SnO(2) nanowires, Under reverse bias, near bandedge emission from the p-GaN:Mg/sapphire leads to the observation of emission peak at around 370 nm.

The novel anti-hyperglycemic effect of Paeoniae radix via the transcriptional suppression of phosphoenopyruvate carboxykinase (PEPCK).

The antidiabetic actions of Paeoniae Radix involve stimulating glucose uptake and reducing glucose absorption. However, the importance of this herb in the transcriptional regulation of hepatic gluconeogenesis has not previously been investigated, although hepatic gluconeogenesis contributes the most to fasting hyperglycemia. Using rats with streptozotocin-induced diabetes and db/db mice, the dose- and time-dependent suppressive effects of the ethanol extract of Paeoniae Radix (PR-Et) on diabetic hyperglycemia and phosphoenopyruvate carboxykinase (PEPCK) transcription are first demonstrated. Second, by employing H4IIE cells, the inhibitory action of PR-Et on both dexamethasone- and 8-bromo-cAMP-induced-PEPCK expression was also confirmed without causing any cytotoxicity. In addition, this inhibitory effect could be sustained for over 24 h with repeated treatment. Most importantly, PR-Et's action was unaffected by either insulin desensitization or palmitate stimulation. Finally, paeonol and paeoniflorin, two well-known constituents in Paeoniae Radix, did not suppress PEPCK expression at testing concentration. In conclusion, it was clearly demonstrated that transcriptional inhibition of gluconeogenesis is one of the important antidiabetic actions of Paeoniae Radix. Future development of this herb as a dietary supplement or drug should bring substantial benefits for the diabetic population.

Identification and characterization of the inward K+ channel in the plasma membrane of Brassica pollen protoplasts.

Patch clamp techniques have been used to identify and characterize the whole-cell currents carried by inward K+ channels in isolated matured pollen protoplasts of Brassica chinensis var. chinensis. The whole-cell inward currents in the isolated pollen protoplasts were activated at hyperpolarized membrane potentials more negative than -100 mV. The magnitudes of the whole-cell inward currents were strongly dependent on the external K+ concentration, and were highly selective for K+ over other monovalent cations. The inward currents were not observed when external K+ was replaced with the same concentration of Cs+ or Na+. The addition of 1 mM or 10 mM Ba2+ in external solutions resulted in 30% or 80% inhibition of the inward currents at -180 mV, respectively. These results demonstrated that the inward K+ currents mainly account for the recorded whole-cell inward currents in Brassica pollen protoplasts. Increase of cytoplasmic Ca2+ concentrations from 10 nM to 30 microM or even 5 mM did not affect the inward K+ currents. Decrease of external Ca2+ concentrations from 10 mM to 1 mM inhibited the inward K+ currents by 25%, while the increase of external Ca2+ from 10 mM to 50 mM almost completely blocked the inward K+ currents. Physiological importance of K+ transport into pollen and its possible regulatory mechanisms are also discussed.

Lacrimal punctal occlusion for the treatment of superior limbic keratoconjunctivitis.

PURPOSE: To test the hypothesis that superior limbic keratoconjunctivitis is caused by insufficient tear supply to the superior keratoconjunctiva. METHODS: We used cautery and sutures to permanently occlude the lacrimal puncta of 11 patients (22 eyes) with superior limbic keratoconjunctivitis for whom topical treatment was ineffective. RESULTS: All 11 patients (22 eyes) responded favorably to lacrimal punctal occlusion. After lacrimal punctal occlusion, rose bengal and fluorescein staining (both on a scale of 0 [no staining] to 9 [complete staining]) were reduced (mean +/- SD, 2.7 +/- 1.6 to 1.1 +/- 1.8 and 1.4 +/- 1.2 to 0.4 +/- 0.8, respectively). Impression cytology disclosed improvement of squamous metaplasia in the superior conjunctiva as well as increased goblet cells in nine of 13 eyes (69%) examined. Subjective symptoms improved in all 22 eyes (100%). CONCLUSIONS: Improvement of local tear deficiency to the superior limbic portion by punctal occlusion was an effective treatment in this small series. Superior limbic keratoconjunctivitis might be caused by the insufficient local tear supply.

Inhibitory effects of some steroidal saponins on human spermatozoa in vitro.

Twenty-nine C-27 steroidal saponins were tested for the inhibitory effects on human spermatozoa in vitro by means of a modified Sander-Cramer method. Twelve of them are responsible for this activity. The structure-activity relationship is discussed.

Direct sub-Tenon's ocular anesthesia for strabismus surgery.

We assessed the safety and effectiveness of sub-Tenon's anesthesia, a local anesthesia, for strabismus surgery. For 15 surgeries, we used anesthesia by sub-Tenon's infusion, followed by direct infusion through a transconjunctival route with a local anesthetic agent. We found the procedure was simple, it minimized complications compared with other techniques and was effective in achieving rapid anesthesia. With the ocular movement remaining, we checked eye position by the alternative cover-uncover test at the end of surgery. We obtained good eye position for all patients. This method leads to good results for strabismus surgery.

Functional cytochrome P4503A isoforms in human embryonic tissues: expression during organogenesis.

Expression of functional cytochrome P450 (CYP) isoforms in human embryonic tissues was explored during organogenesis (days 50-60 of gestation) with substrate probes, inhibitor probes, and immunoprobes and by reverse transcription-polymerase chain reaction (PCR), cloning, and sequencing. Evidence was obtained for the presence of relatively high levels of one or more functional CYP3A isoforms in embryonic livers. This was manifested as relatively extensive hydroxylation of (R)-warfarin at carbon 10 and as triacetyloleandomycin-inhibited O-debenzylation of benzyloxyresorufin when human embryonic hepatic microsomal fractions were used as enzyme sources. Immunoblots with anti-CYP3A4 antibody exhibited a strong signal in embryonic hepatic tissues but, in contrast, indicated very low or negligible CYP3A levels in human embryonic lung, kidney, heart, adrenal, and brain tissues. To explore expression of individual members of the CYP3A subfamily in human embryonic hepatic tissues at this early gestational stage, CYP3A cDNA was generated by reverse transcription, amplified by PCR, cloned, and sequenced. Oligonucleotide primers used for PCR were designed to flank target sequences unique to CYP3A but also common to all human CYP3A subfamily members for which GenBank nucleotide sequence information was available (CYP3A3, CYP3A4, CYP3A5, CYP3A5P, and CYP3A7). Sequencing data indicated that plasmids in 58 of 59 recombinant positive colonies contained an insert with a sequence identical to that present in CYP3A7 cDNA and the plasmid of only one colony contained an insert with a sequence identical to that present in CYP3A5 cDNA. No evidence was found for expression of CYP3A3 or CYP3A4. Thus, during organogenesis, human embryonic hepatic tissues express primarily CYP3A7 and are capable of significant CYP3A7-catalyzed xenobiotic monooxygenation during this very early stage of gestation.

FMRF-NH2-like peptide is deficient in the pituitary gland of the Brattleboro rat.

Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-NH2 (F-8-F-NH2), isolated from bovine brain, is an FMRF-NH2-like peptide with morphine-modulating activity. In the rat, F-8-F-NH2 immunoreactivity (IR) is highly localized in the neurohypophysis. In this study, F-8-F-NH2-IR was studied in the hypothalamo-neurohypophyseal system of an Arg8-vasopressin (AVP)-deficient animal, the Brattleboro (DI) rat, and the normal control Long-Evans (LE) strain. F-8-F-NH2-IR in the DI pituitary is below the level of detection in contrast to that in the LE (0.50 +/- 0.04 pmol/gland). Neuropeptide Y (NPY) levels are increased two-fold in the DI pituitary while AVP levels are below detection. The content of F-8-F-NH2-IR in the hypothalami and spinal cords of DI and LE rats is not statistically different, suggesting that the absence of F-8-F-NH2-IR in the Brattleboro pituitary is not due to a genetic defect in F-8-F-NH2 biosynthesis. The results of this study raise the question whether AVP could be involved in the regulation of F-8-F-NH2 immunoreactivity in the neurohypophysis.

Clinical study of syncope during acupuncture treatment.

From August 1988 to April 1989, we observed 52 patients who developed so-called 'needle fainting' (or what the Chinese call 'Yun-Cheng' phenomenon) 55 times among a total sample of 28,285 procedures of acupuncture therapy at the Center for Traditional Medicine of Veterans General Hospital in Taipei. Of these syncopal patients, 35 were male and 17 were female. Their mean age was 45 years (with a range of 11 to 72 years). All patients were in an upright position when needle fainting occurred. Their usual manifestations were pallor, cold sweating, nausea, and bradycardia. They all recovered soon after lying down; no one developed a complete loss of consciousness. No mortality was noted. When comparing the patients who experienced syncope during their first visit to our Clinic (Group I, n = 27) with the patients who experienced syncope in a follow-up treatment (Group II, n = 25; 3 patients had 2 episodes in sequential treatments), we found a significantly higher incidence of needle fainting (p less than 0.0001) in Group I patients (27 out of 2,855 or 0.94%) than in Group II patients (28 out of 25,430 or 0.11%). The mean age of Group I patients (39 +/- 15.4 years) was significantly less than that of Group II patients (51.6 +/- 18.0 years) (p less than 0.001). The coexistence of other medical problems was significantly higher in Group II patients (72%) than in Group I patients (18.5%) (p less than 0.0001).

Release of high molecular weight forms of met5-enkephalin-arg6-gly7-leu8 from rat brain.

The release of various molecular weight forms of met5-enk-arg6-gly7-leu8 (MERGL) from slices prepared from rat medulla-pons and hypothalamus was studied. The release of MERGL-IR from both medulla-pons and hypothalamic slices was elevated four- to ten-fold in response to potassium depolarization. Gel of 8 to 10 kilodalton MERGL-IR peptide(s), as well as MERGL itself, were released in response to stimulation with 50 mM K+.

Reevaluation of 5-fluorouracil as a single therapeutic agent for gestational trophoblastic neoplasms.

Large dosage of 5-fluorouracil given by slow intravenous infusion has proved to be very effective in the treatment of gestational trophoblastic neoplasms. From 1965 through 1975, 5-fluorouracil was used as a single chemotherapeutic agent in 173 cases of invasive mole and 139 cases of choriocarcinoma. Complete remission was achieved in 84.9% of cases of invasive mole and in 59.3% of cases of choriocarcinoma when 5-fluorouracil was used as the initial treatment. Seven recurrences with three deaths occurred during follow-up in 216 patients who had achieved complete remission, providing a recurrence rate of 3.2% and recurrence death rate of 1.4%. All of the survivors were followed up for more than 5 years and 85.6% for more than 10 years. Toxicity of 5-fluorouracil was milder and less frequent than that of 6-mercaptopurine or methotrexate. The toxic reaction specific to 5-fluorouracil was diarrhea, which can result in pseudomembranous colitis if improperly treated. One of the two toxic deaths was due to this complication.